In Vitro Evidence Limitations

In vitro studies are useful for mechanism — they are not evidence of clinical effect.

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What in vitro studies establish

That a compound can bind a given receptor.
That a measurable downstream change occurs in a defined cell line.
That the effect is dose-dependent in a defined concentration range.

What in vitro studies do not establish

That the effect occurs in vivo.
That the concentration used is achievable in human tissue.
That the cell line resembles the relevant human tissue.
That the effect translates into a clinical outcome.

The concentration-relevance problem

Cell-culture experiments often use compound concentrations that would never be achieved in human plasma or tissue. A finding at 100 µM in a culture dish is mechanistically interesting but says little about what will happen at the 1–100 nM concentrations that real-world dosing might produce.

Cell-line drift and selection

Immortalised cell lines accumulate genetic and phenotypic changes over time. A result in HEK293 or HEK293T cells may not generalise to primary cells of the same tissue type.

Why we grade in vitro evidence at D

On our evidence-grading methodology, a claim supported only by in vitro evidence sits at Grade D. The claim is mechanistically plausible but is not a human-relevant finding without further work. See: evidence grading.